Project Summary The microbiome and the metabolome are intricately linked with host immunity and are emerging as critical regulators of vaccine-induced immune responses. Despite their importance, we lack a fundamental understanding of their influence ? both positive and negative on HIV vaccine outcomes. Bridging this gap will complement the significant efforts directed at immunogen/adjuvant design and advance the development of an HIV vaccine that is both effective and safe. The main objective of this R03 is to delineate the role of the microbiome and metabolome in accumulation of HIV target CD4 T cells in the female genital tract (FGT) in order to understand their role in subverting vaccine efficacy. These studies will be conducted under the auspices of a parent K01-funded vaccine study evaluating immunogencity and efficacy of a DNA-Prime/Protein boost vaccine platform in rhesus macaques. In Aim 1 of this research project, we will test the hypothesis that recruitment of HIV target CD4 T cells to the lower FGT over the course of vaccination is dictated by specific vaginal bacterial communities. In Aim 2, we will determine whether distinct metabolite profiles are strongly associated with the frequency and phenotype of HIV target cells in the FGT. The ultimate goal of this R03 proposal is to determine whether a cervicovaginal microbial/metabolite composition that predisposes to vaccine induced infiltration of HIV target CD4 T cells to the FGT results in vaccine failure. By integrating cutting-edge microbiome and metabolome analysis into scheduled collection of samples alongside a comprehensive evaluation of immune responses and vaccine efficacy, the R03 will leverage existing resources to not only break new ground and open avenues for further exploration but also better inform vaccine outcomes of the parent study. The findings will provide critical preliminary data for an R01 proposal to elucidate the mechanisms by which microbiota influence HIV vaccine outcomes. The strong environment of the CNPRC and CCM, robust genomic, metabolomic, bioinformatics, and statistical support provided by UC Davis Genome Center, the strong and complementary expertise of the collaborators combined with the PIs expertise in T cell immunology, preclinical vaccine studies, micobiota analysis, and metabolism will facilitate the successful execution of the proposed studies within the 2-year timeline.